title: Neuronal swelling and surface area regulation: elevated intracellular calcium is not a requirement.
abstract: Neurons are mechanically robust. During prolonged swelling, molluscan neurons can triple their apparent membrane area. They gain surface area and capacitance independent of extracellular Ca concentration ([Ca]e), but it is unknown if an increase in intracellular Ca concentration ([Ca]i) is necessary. If Ca for stimulating exocytosis is unnecessary, it is possible that swelling-induced membrane tension changes directly trigger surface area readjustments. If, however, Ca-mediated but not tension-mediated membrane recruitment is responsible for surface area increases, swelling neurons should sustain elevated levels of [Ca]i. The purpose of this investigation is to determine if the [Ca]i in swelling neurons attains levels high enough to promote exocytosis and if any such increase is required. Lymnaea neurons were loaded with the Ca concentration indicator fura 2. Calibration was performed in situ using 4-bromo-A-23187 and Ca-ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), with free Ca concentration ranging from 0 to 5 microM. Swelling perturbations (medium osmolarity reduced to 25% for 5 min) were done at either a standard [Ca]e or very low [Ca]e level (0.9 mM or 0.13 microM, respectively). In neither case did the [Ca]i increase to levels that drive exocytosis. We also monitored osmomechanically driven membrane dynamics [swelling, then formation and reversal of vacuole-like dilations (VLDs)] with the [Ca]i clamped below 40 nM via 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). [Ca]i did not change with swelling, and VLD behavior was unaffected, consistent with tension-driven, [Ca]i-independent surface area adjustments. In addition, neurons with [Ca]i clamped at 0.1 microM via an ionophore could produce VLDs. We conclude that, under mechanical stress, neuronal membranes are compliant by virtue of surface area regulatory adjustments that operate independent of [Ca]i. The findings support the hypothesis that plasma membrane area is regulated in part by membrane tension.
authors.affiliation: Department of Biology, University of Ottawa, Ontario, Canada.
authors.forename: T L
authors.initials: TL
authors.lastname: Herring
authors.forename: I M
authors.initials: IM
authors.lastname: Slotin
authors.forename: J M
authors.initials: JM
authors.lastname: Baltz
authors.forename: C E
authors.initials: CE
authors.lastname: Morris
mesh_terms: D000818:Animals; D000001:Calcimycin / Q000031:analogs & derivatives / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D004533:Egtazic Acid / Q000494:pharmacology; D017952:Ganglia, Invertebrate / Q000166:cytology / Q000502:physiology*; D066298:In Vitro Techniques; D008195:Lymnaea; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D013499:Surface Properties; D014617:Vacuoles / Q000502:physiology / Q000648:ultrastructure
chemical_list: D000001:Calcimycin; D004533:Egtazic Acid; C033170:4-bromo-A-23187; D002118:Calcium